Corynebacterium kutscheri is a pathogen which produces pseudotuberculosis in laboratory animals. The infection commonly exists undetected in a putative latent form which becomes active when infected animals are subjected to resistance-lowering stress. Once C. kutscheri is introduced into a colony of laboratory animals, the pathogen is difficult to eradicate. The pathogenicity for the mouse and rat is well documented. Since infected animals cannot be used for experimentation, an effective method of rapidly evaluating laboratory animals for C. kutscheri infection would be extremely helpful.
The putative latent form of C. kutscheri infection is difficult to detect. Methods of detecting unapparent infections includes provocation testing with cortisone acetate, immunological testing, and serological testing by anamnesic response. Provocation tests are effective in detecting silent contamination, but result in propagation of infective agents. Diagnostic results are obtained in 7 to 10 days. Immunological tests are not completely effective because recently infected animals require 10 to 14 days to produce detectable amounts of antibody. Immunological tests also involve cross-reactivity problems.
Hybridization-probes are known in the field of molecular biology. U.S. Pat. No. 4,358,535, issued to Falkow on Nov. 9, 1982, discloses a process for detecting DNA sequences of pathogens of interest in raw clinical samples. Samples are placed onto a solid support and deposited microbes are treated to release DNA. The DNA is complexed onto the support and denatured. A labeled polynucleotide probe specific for a DNA sequence characteristic of a pathogen is contacted to the support under hybridization conditions. Hybridization of pathogenic DNA is detected by means of the probe label.
Wirth and Pratt, Proc. Natl. Acad. Sci., 79: 6999 to 7003 (1982) disclose a method for detecting kinetoplast DNA (kDNA) of Leishmania species in cutaneous lesions. Blots are prepared by touching excised lesions on nitrocellulose filters. Species-related minicircles of kDNA are isolated from cultured Leishmania promastigotes and labeled radioisotopically. The nitrocellulose filter is hybridized with the labeled kDNA. Hybridization is detected on film.
U.S. Pat. No. 4,446,237, issued to Berninger on May 1, 1984, discloses a method for detecting viral DNA in acellular biological fluid. Solutions of denatured DNA isolated from acellular biological fluid are applied to a solid support. Viral DNA is denatured and labeled radioisotopically. The labeled DNA is contacted to the support under hybridization conditions. Hybridization of viral DNA is detected by means of the radioisotope label.
The following are representative of other typical DNA probe references. Chou and Merigan, New England Journal of Medicine, 308: 921 (1983) disclose the detection of cytomegalovirus DNA in clinical urine specimens after immobilization on nitrocellulose filters and hybridization with a radioactively labeled, cloned fragment of cytomegalovirus DNA. Spector et al., J. Infectious Diseases, 150: 121 (1984) disclose a diagnostic assay for the detection of human cytomegalovirus (HCMV) DNA in clinical specimens with .sup.32 P-labeled cloned fragments at HCMV DNA.